PROJECT ABSTRACT This application is for a Mentored Patient-Oriented Research Career Development Award entitled ?Microbiome-based diagnosis of pneumonia in the acute respiratory distress syndrome?. I am a Pulmonary and Critical Care Medicine Fellow at the University of Pittsburgh and I require additional training to develop expertise as a translational researcher in the microbiome of critical illness. The central objective of my research is to utilize emerging microbial DNA sequencing techniques for faster and more accurate diagnosis of pneumonia in critically-ill patients. The focus of my research is the acute respiratory distress syndrome (ARDS), a serious form of inflammatory lung injury often caused by pneumonia and accounting for 10% of intensive care unit admissions in the US with 40% mortality. With diagnostic limitations of the current standard- of-care of microbial cultures, patients often receive empiric, broad-spectrum antibiotics that can be inappropriate or inadequate for their individual needs, and also promote antibiotic resistance ? a major public health threat. My preliminary data demonstrate feasibility and proof-of-concept that sequencing of microbial DNA directly from patient samples can lead to faster and more accurate diagnosis of pneumonia compared to cultures. The aims of the study are to develop sequencing-based definitions for pneumonia in (1) culture- positive and (2) culture-negative patients by integrating microbiome and host response data, and then 3) to assess the anticipated impact of sequencing on antibiotic selection. I will execute a prospective cohort study in patients with ARDS and use modern culture-independent sequencing techniques and computational biology methods to study the lung microbiome in relation to host-responses and microbiologic cultures. The proposed sequencing-based approach has the potential to improve accuracy of pneumonia diagnosis (capturing both cultivable and non-cultivable organisms) and to provide information to clinicians much faster than cultures. Thus, my proposal addresses a high priority issue for development of innovative diagnostics of infections to improve antibiotic usage and provides a unique translational training opportunity. I propose a research- intensive period of career development with hands-on and didactic training in advanced sequencing techniques, bioinformatics, computational biology and clinical trial design and implementation. The work will be conducted within the Division of Pulmonary, Allergy, and Critical Care Medicine at the University of Pittsburgh, which has an outstanding record of training physician-scientists and a highly-developed infrastructure for the conduct of translational studies. I am supported by a committed mentoring partnership with convergent expertise in microbiome and critical care research, and a diverse Mentoring Advisory Committee. With support from the K23 Mentored Career Development Award, I will develop an independent project examining sequencing-based pneumonia diagnostics for an R01 proposal over the next five years.